In the Xenopus oocyte expression system, the NMDA receptor subunit NR1 has been shown to be inhibited by alcohol and organic solvent anesthetics. Amino acid residues within the TM3 domain of the receptor are important for this inhibition. A change of a phenylalanine to alanine at position 639 within TM3 eliminates inhibition by some anesthetics and reduces the efficacy of alcohol inhibition in oocytes. If this inhibition of the NMDA receptor contributes significantly to behavioral effects of alcohol, then one would predict that an animal carrying this mutation will be less sensitive to alcohol. Establishing this connection can be labor intensive in the mouse model system. Drosophila provides an attractive alternative as a high-throughput system for mutant analysis of genes since knock-in mutants can be produced rapidly and at a fraction of the cost of mouse transgenics. The NR1 gene is a good candidate to test this approach for correlating Xenopus structure-function studies to animal behavior for the following reasons: the fly NR1 TM3 domain is 95% identical to the mouse TM3 domain, this receptor is involved in similar processes in both flies and mammals (e.g. learning and memory) and finally, flies and mammals have similar responses to alcohol and anesthetics. We will use Drosophila gene knock-in technology to generate flies carrying the F639A mutation within the NR1 receptor gene. These flies will be examined to determine if the mutation reduces the sensitivity to sedation with alcohol and organic solvent anesthetics. Simultaneously, we will also determine if the mutation affects the capacity to acquire behavioral tolerance to sedation with these drugs. Following this pilot study, we will extend this work to more broadly correlate oocyte electrophysiological data on the effects of alcohol and anesthetics to whole animal behavioral data in flies.